Cloning and Characterization of 4EHP, a Novel Mammalian eIF4E-related Cap-binding Protein

Rom, Eran; Kim, Hyung Chan; Gingras, Anne‐Claude; Marcotrigiano, Joseph; Favre, Daniel; Olsen, Henrik S.; Burley, S.K.; Sonenberg, Nahum

doi:10.1074/jbc.273.21.13104

Cited by 132 publications

(159 citation statements)

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“…We performed a complementation assay using tethered λN-CNOT1 in the presence of WT 4EHP or the 4EHP W124A mutant, which is incapable of binding to the cap (19). Transient transfection of shRNA-resistant 3xFlag-4EHP in 4EHP-knockdown cells restored λN-CNOT1-mediated translation repression (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Cap-binding protein 4EHP effects translation silencing by microRNAs

Chapat

Jafarnejad

Matta‐Camacho

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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111

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MicroRNAs (miRNAs) play critical roles in a broad variety of biological processes by inhibiting translation initiation and by destabilizing target mRNAs. The CCR4-NOT complex effects miRNA-mediated silencing, at least in part through interactions with 4E-T (eIF4E transporter) protein, but the precise mechanism is unknown. Here we show that the cap-binding eIF4E-homologous protein 4EHP is an integral component of the miRNA-mediated silencing machinery. We demonstrate that the cap-binding activity of 4EHP contributes to the translational silencing by miRNAs through the CCR4-NOT complex. Our results show that 4EHP competes with eIF4E for binding to 4E-T, and this interaction increases the affinity of 4EHP for the cap. We propose a model wherein the 4E-T/4EHP interaction engenders a closed-loop mRNA conformation that blocks translational initiation of miRNA targets.M icroRNAs (miRNAs) are short, ∼22-nucleotide noncoding RNAs that affect gene expression in most eukaryotes. miRNAs mediate posttranscriptional silencing by guiding the miRNA-induced silencing complex (miRISC), an assembly of Argonautes and GW182/TNRC6 proteins, to target mRNAs. Target recognition initiates a succession of events: mRNA translational repression, deadenylation, and mRNA decay (1). miRNAs impair the function of eIF4F, a three-subunit complex composed of eIF4E, the m 7 GTP (cap)-interacting factor; eIF4G, a scaffolding protein; and eIF4A, a DEAD-box RNA helicase (2-5). The silencing activity of miRISC is mediated by the CCR4-NOT deadenylase complex through the scaffolding subunit, CNOT1 (6-8). CNOT1 recruits the DDX6 and 4E-T (eIF4E transporter, also known as EIF4ENIF1) proteins, which are important for miRNA-mediated silencing (9-16). The 4E-T protein is a conserved eIF4E-binding protein, which directly binds to the dorsal surface of eIF4E through its canonical eIF4E-binding YX 4 LL (Y 30 TKEELL) motif and impairs the eIF4E/eIF4G interaction and translation initiation (17). The 4E-T protein also facilitates the decay of CCR4-NOT-targeted mRNAs by linking the 3′-terminal mRNA decay machinery to the cap via its interaction with eIF4E (13).In mammals, eIF4E is the best-studied member of a family of proteins composed of eIF4E (eIF4E1), 4EHP (4E-homologous protein; eIF4E2), and eIF4E3. The 4EHP and eIF4E proteins share 28% sequence identity (18,19). The 4EHP protein is ubiquitously expressed, and it is 5-10 times less abundant than eIF4E in a number of mammalian cell lines (18)(19)(20). Like eIF4E, 4EHP binds to 4E-T, but in sharp contrast to eIF4E, it does not associate with eIF4G (18, 21). The 4EHP protein has a 30-to 100-fold weaker affinity for the cap than eIF4E due to a twoamino acid substitution in its cap-binding pocket (22).The 4EHP protein has primarily been documented as a translation repressor. In the Drosophila embryo, 4EHP associates with the RNA binding protein Bicoid to repress caudal mRNA translation (23). Similarly, 4EHP also represses the hunchback mRNA by binding to the nanos repressive element complex, which consists of nanos...

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Section: Resultsmentioning

confidence: 99%

“…The 4EHP and eIF4E proteins share 28% sequence identity (18,19). The 4EHP protein is ubiquitously expressed, and it is 5-10 times less abundant than eIF4E in a number of mammalian cell lines (18)(19)(20). Like eIF4E, 4EHP binds to 4E-T, but in sharp contrast to eIF4E, it does not associate with eIF4G (18, 21).…”

mentioning

confidence: 99%

Cap-binding protein 4EHP effects translation silencing by microRNAs

Chapat

Jafarnejad

Matta‐Camacho

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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111

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“…3,4 eIF4E2 was first characterized as a translation inhibitor due to its ability to bind the 5′cap but not eIF4G, which is required to recruit ribosomes. 5 However, in hypoxia eIF4E2 interacts with eIF4G3 4 and initiates the translation of mRNAs containing 3′UTR RNA hypoxia response elements bound by hypoxia-inducible factor-2α. 3 eIF4E2 is required for tumor progression 6 and is part of a metastatic gene signature.…”

Section: Introductionmentioning

confidence: 99%

Hypoxia activates cadherin-22 synthesis via eIF4E2 to drive cancer cell migration, invasion and adhesion

et al. 2017

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Hypoxia is a driver of cell movement in processes such as development and tumor progression. The cellular response to hypoxia involves a transcriptional program mediated by hypoxiainducible factors, but translational control has emerged as a significant contributor. In this study, we demonstrate that a cell-cell adhesion molecule, cadherin-22, is upregulated in hypoxia via mTORC1-independent translational control by the initiation factor eIF4E2. We identify new functions of cadherin-22 as a hypoxia-specific cell-surface molecule involved in cancer cell migration, invasion and adhesion. Silencing eIF4E2 or cadherin-22 significantly impaired MDA-MB-231 breast carcinoma and U87MG glioblastoma cell migration and invasion only in hypoxia, while reintroduction of the respective exogenous gene restored the normal phenotype. Cadherin-22 was evenly distributed throughout spheroids and required for their formation and support of a hypoxic core. Conversely, E-cadherin translation was repressed by hypoxia and only expressed in the oxygenated cells of U87MG spheroids. Furthermore, immunofluorescence on paraffinembedded human tissue from 40 glioma and 40 invasive ductal breast carcinoma patient specimens revealed that cadherin-22 expression colocalized with areas of hypoxia and significantly correlated with tumor grade and progression-free survival or stage and tumor size, respectively. This study broadens our understanding of tumor progression and metastasis by highlighting cadherin-22 as a potential new target of cancer therapy to disable hypoxic cancer cell motility and adhesion.

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“…eIF4E homologs have been identified in most organisms (except for Saccharomyces cervisiae). In mammals, there are three members of the eIF4E family: eIF4E (eIF4E-1), 4EHP (eIF4E-2), and eIF4E-3 (Rom et al 1998;Joshi et al 2004). Based on sequence alignments, h4EHP, together with nCBP from plants (Ruud et al 1998), IF4E-4 from Caenorbiditis elegans and d4EHP (deIF4E-8) from Drosophila melanogaster are members of the eIF4E-2 class (Joshi et al 2005).…”

Section: Introductionmentioning

confidence: 99%

Weak binding affinity of human 4EHP for mRNA cap analogs

Zuberek¹,

Kubacka

Jabłonowska³

et al. 2007

RNA

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Ribosome recruitment to the majority of eukaryotic mRNAs is facilitated by the interaction of the cap binding protein, eIF4E, with the mRNA 59 cap structure. eIF4E stimulates translation through its interaction with a scaffolding protein, eIF4G, which helps to recruit the ribosome. Metazoans also contain a homolog of eIF4E, termed 4EHP, which binds the cap structure, but not eIF4G, and thus cannot stimulate translation, but it instead inhibits the translation of only one known, and possibly subset mRNAs. To understand why 4EHP does not inhibit general translation, we studied the binding affinity of 4EHP for cap analogs using two methods: fluorescence titration and stopped-flow measurements. We show that 4EHP binds cap analogs m 7 GpppG and m 7 GTP with 30 and 100 lower affinity than eIF4E. Thus, 4EHP cannot compete with eIF4E for binding to the cap structure of most mRNAs.

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Cloning and Characterization of 4EHP, a Novel Mammalian eIF4E-related Cap-binding Protein

Cited by 132 publications

References 59 publications

Cap-binding protein 4EHP effects translation silencing by microRNAs

Cap-binding protein 4EHP effects translation silencing by microRNAs

Hypoxia activates cadherin-22 synthesis via eIF4E2 to drive cancer cell migration, invasion and adhesion

Weak binding affinity of human 4EHP for mRNA cap analogs

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